A Single-Cell RNA-Seq-Based Approach for Genome-Wide Identification of Cell Essential Genes.

Identification of genes essential for structure, function, and survival of a cell type is critical for understanding of the underlying mechanisms. Unfortunately, there is no efficient way to identify such genes. Studies by single-cell RNA sequencing have shown that gene expressions of single cells of the same type are highly heterogeneous. We therefore speculate that the genes expressed in all individual cells of the same type are essential for the cell type, including the housekeeping genes and cell type-specific essential genes. Based on this rationale, we design a high-throughput approach to identify podocyte essential genes. In this approach, mouse podocytes are subjected to ultra-deep single-cell RNA-seq, and the genes expressed in all single podocytes are sorted out and considered as the candidates of podocyte essential genes. The essentiality of these genes for podocytes is assessed by bioinformatics, cross-species conserved expression, association with injury/disease, inclusion of known essential genes, and experimental validation. By comparison with the essential genes of other cell types, podocyte-specific essential genes can be distinguished. This approach applies to any cell types. In this chapter, we describe the approach and detailed methods.